Method and apparatus for measuring osmotic pressure

ABSTRACT

A method and apparatus are provided for the measuring osmotic pressure of a solution. A sample of a solution is placed into a sample cell above a membrane supported in the sample cell and the sample cell is sealed. Gas at an elevated pressure is introduced into the sample cell to drive a quantity of dialyzate through the membrane and into a transparent dialyzate exit tube adapted to receive the dialyzate which passes through the membrane. The elevated pressure of the gas is varied through a pressure regulator to yield a substantially stationary dialyzate meniscus in the transparent dialyzate exit tube. The pressure in the sample cell is measured by reading a pressure gauge showing the elevated pressure of the gas that yielded the substantially stationary dialyzate meniscus, the elevated pressure of the gas being substantially equivalent to the osmotic pressure of the sample.

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] The present application is related to U.S. Application Ser. No.(Docket No 80378), filed Herewith, by Dr. David C. Boris, and entitled,“METHOD FOR MEASURING CHANGES IN OSMOTIC PRESSURE,” and to U.S.Application Ser. No. (Docket No. 80379), filed Herewith, by Dr. David C.Boris, and entitled, “METHOD FOR DETERMINING THE CONCENTRATION OF ASOLUTION HAVING A PREDETERMINED OSMOTIC PRESSURE AND/OR GENERATING ASOLUTION WITH A PREDETERMINED OSMOTIC PRESSURE.”

FIELD OF THE INVENTION

[0002] The present invention relates generally to osmometry and, moreparticularly, to measuring the osmotic pressure of complex solutionsincluding, but not limited to, emulsions, dispersions and charged anduncharged polymer solutions.

BACKGROUND OF THE INVENTION

[0003] A variety of methods and devices are known for measuring theosmotic pressure exerted by solvent molecules diffusing through assemipermeable membrane. Commercially available devices for measuring theosmolality of solutions via membrane osmometry include the 4400 seriescolloid osmometers made by WesCor (Logan, Utah) and the type 1.00 Knaurmembrane osmometers. Other devices for measuring the osmolality ofsolutions through vapor pressure osmometry are commercially available.An example of this type of device is the VPO model 070 made by UIC Inc.(Joliet, Ill.). Still another known osmometer device operates bymeasuring freezing point depression. An example of this type of deviceis the Precision Systems Inc. (Natick, Mass.) Osmette XL product line.Other known osmometer variants include isopiestic vapor equilibriumosmometers and submerged dialysis bag osmometers. In addition,osmolality has been determined by measuring boiling point elevation. Thevapor pressure osmometer, boiling point elevation, freezing pointdepression, and isopiestic methods measure the oncotic pressure (theosmotic pressure exerted by colloid in the solution) of the solution,that is, the osmolality of the solution including the contribution oflow molecular weight components such as salts. The present invention isdirected to methods for measuring the osmotic pressure of solutions,excluding the contribution of small molecules.

[0004] Membrane osmometers and the dialysis bag techniques measure thesolution equilibration across a semi-permeable membrane, thus excludingthe direct contribution of small permeable molecules. One example of amembrane osmometer is taught in U.S. Pat. No. 4,150,564, titled“OSMOMETER FOR COLLOID OSMOMETRY,” by Wayne K. Barlow, et al., Apr. 24,1979. The present invention is an improvement upon existing membraneosmometer designs, but relies on the same basic principle ofestablishing an equilibrium across a semi-permeable membrane. Typicalmembrane osmometers employ pressure transducer technology to directlymeasure the evolved osmotic pressure difference between a reference celland the sample solution across a semi-permeable membrane. In the presentinvention transducer technology is not required for measuring theosmotic pressure. Typical commercial osmometers are designed to minimizethe sample volumes by introducing the sample into a meandering channelabove the semi-permeable membrane that maximizes surface contact whileminimizing sample volume. This meandering channel geometry is eliminatedin the present invention because it limits the usefulness of commercialosmometers to low viscosity, non-fouling solutions and is particularlyunsuited for complex solutions (dispersions and emulsions).

[0005] Dialysis bag techniques, (Essafi, W. Structure Despolyelectrolytes Fortement Charges, PhD thesis, Universite Pierre etMarie Curie, Paris, 1996) involve filling a semi-permeable dialysis bagwith the sample solutions of unknown osmotic pressure and immersing itin a large volume of solution of known osmotic pressure. The samplechanges concentration until an osmotic equilibrium is established. Thenthe sample is removed and its concentration at that known osmoticpressure is determined using other techniques (spectrophotometrically orgravimetrically). The present invention may be used in a mode ofoperation similar to this. In this mode of operation the sample solutionis allowed to equilibrate to a known imposed air pressure. Then theconcentrated sample is removed and the concentration determinedseparately. However, this method of operation is not the optimal nor isit the preferred embodiment of the method of the present invention.

[0006] The prior art also suggests use of polymer solution s of knownosmotic pressure to have as reference solutions (rather than solvent)using commercial osmometers or other direct force measurement techniquesto allow for measurement of higher osmotic pressure solutions, (Rau,Donald C.; Parsegian, V. Adrian. Direct Measurement OfTemperature-Dependent Solvation Forces Between DNA Double Helixes.Biophys. J. (1992), 61(1), 260-71; Sidorova, Nina Y.; Rau, Donald C.Removing Water From An EcoRI-noncognate DNA Complex With Osmotic Stress.J. Biomol. Struct. Dyn. (1999), 17(1), 19-31). This method worksreasonably well for extending the pressure range available on commercialosmometers but does not properly treat the Donnan equilibriumestablished for charged species thereby potentially leading to erroneousresults with charged polymers. This method of extending the pressurerange is not needed in the operation of the present invention as thereference solution is simply dialyzate.

[0007] The present invention capitalizes upon advances in stirred celldialysis chamber technology to improve osmometer design. Stirred celldialysis chambers are available from, for example, Amicon, (Beverly,Mass.). The principal use of the dialysis chamber in the prior art is toeither concentrate a sample solution or to remove small moleculeimpurities by exhaustive flushing with pure solvent. The stirred cell isdesigned to be dismantled easily for cleaning. It has a magneticstirring rod suspended above the membrane to keep the solution wellstirred and to sweep clean the surface of the membrane. It can withstandmore than 75 psi of external pressure, far exceeding the pressuremeasurable in conventional membrane osmometers (˜1-3 psi).

SUMMARY OF THE INVENTION

[0008] It is therefore an object of the present invention to provide anapparatus for measuring osmotic pressure which can be easily dismantledand cleaned so that complex fluids (the emulsions, dispersions, etc.)which would foul traditional meandering channel membrane osmometers canbe measured.

[0009] A further object of the present invention is to provide anapparatus for measuring osmotic pressure which includes a stirrer thatsweeps the surface of the membrane continually thereby reducing surfacefouling and removing bubbles that hamper the accuracy and reliability ofprior art osmometers when used to measure complex fluids.

[0010] Yet another object of the present invention is to provide anosmometer which keeps the sample well mixed thereby avoiding particlesettling and the development of surface concentration gradients.

[0011] It is a further object of the present invention to pro vide amethod and apparatus for measuring osmotic pressure which can make suchmeasurements more quickly than then conventional prior art osmometers.

[0012] It is yet another object of the present invention to provide amethod and apparatus for measuring osmotic pressure which does notrequire multiple flushes of each sample solution before concentrationsstop changing (as a result of dilution of the sample reservoir due toequilibration with the reference reservoir) and the results arereproducible.

[0013] Still another object of the present invention is to provide amethod and apparatus which can be used for measuring osmotic pressure ofhigh viscosity solutions (above about 100 cp).

[0014] The foregoing and numerous other features, objects and advantagesof the present invention will become readily apparent upon reviewing thedetailed description, claims and drawings set forth herein. Thesefeatures, objects and advantages are accomplished by providing a samplecell with a removable pressurizing lid through which a sample solutionmay be introduced into the chamber of the sample cell. The sample isintroduced above a membrane residing in the sample cell with ameandering dialyzate cell positioned below the membrane. Pressurized gasis introduced into the sample cell via a pressure regulator. Once thesample is forced through the membrane and dialyzate begins to emergethrough a transparent dialyzate tube from the meandering dialyzate cell,the pressure through the pressure regulator is reduced until flowceases. A magnetic stirrer is used to continually sweep the surface ofthe membrane. The primary purposes of the meandering dialyzate cell areto provide a structure for collecting dialyzate with maximum surfacearea contact with the membrane (for fast equilibration with the samplecell) which minimizes the volume of dialyzate relative to the volume ofsample in the sample cell. However, those skilled in the art willrecognize that the present invention can be practiced without ameandering dialyzate cell. Instead, the sample cell can easily beprovided with a support structure other than a meandering dialyzate cellto support the membrane therein. Similarly, the sample cell can also beprovided with a different dialyzate collector (e.g. a funnel). Thesupport structure for the membrane and the dialyzate collector do nothave to be integrated as is the case with a meandering dialyzate cell.However, these alternative arrangements would likely (although notnecessarily) increase the volume of dialyzate needed before a meniscusis achieved in the transparent dialyzate exit tube.

[0015] The use of a pressure regulator and an accurate pressure gaugeallow for precise control and measurement of the applied air pressure onthe sample. By systematically varying the applied air pressure until thevisually observed liquid meniscus of the dialyzate in the dialyzate tubeis stationary, the osmotic pressure of the solution across the dialysismembrane is exactly balanced by the imposed air pressure. Further, themethod and apparatus of the present invention allow for the progressiveadding of known amounts of diluent to the sample solution and mixing thenew dilutions directly in the sample cell. This is achieved practicallyby simply placing the entire cell on an accurate balance and weighingthe amount of added solution before each osmotic measurement.Calibration of the apparatus of the present invention does not requireequilibration of a sample. Rather, calibration is a simple and fastprocedure based on accurately measuring the air-line pressure. This isin marked contrast to the difficulty of repeatedly calibratingtransducers and properly seating membranes in prior art membraneosmometers. This leads to a significant advantage in the amount of timerequired to operate the apparatus.

[0016] For improved accuracy the height of both the dialyzate liquidcolumn in the dialyzate tube and the sample reservoir are recorded andused to correct for the known hydrostatic pressure difference. Theentire apparatus is placed on a hot plate for temperature control. Thismethod is very fast, approximately 4-50 times faster than other methods.Because of the speed of measurement of the present invention, theosmotic pressure of chemical reactions can be monitored as a function oftime or temperature in the sample cell. This feature potentially opensnew areas of research for instance in medicine, or polymer chemistrywhere the appearance or disappearance of reactants is accompanied by anosmotic pressure change.

BRIEF DESCRIPTION OF THE DRAWINGS

[0017]FIG. 1 is a schematic cross-sectional view of the osmometer deviceof the present invention;

[0018]FIG. 2 is a top plan view of the meandering dialyzate cell;

[0019]FIG. 3 is a perspective view of the frame in which the osmometerdevice of FIG. 1 resides during operation;

[0020]FIG. 4 is a graph showing a comparison of osmotic pressure versusconcentration data on Polystyrene Sulfonate obtained using the presentinvention and compared directly to data using a conventional prior artmembrane osmometer; and

[0021]FIG. 5 plots osmotic pressure data from three complex solutionsnot amenable to conventional prior art osmometry techniques including asilver halide containing gelatin photographic dispersion, a carbonparticle slurry and a high viscosity charged polymer solution.

DESCRIPTION OF THE PREFERRED EMBODIMENT

[0022] Turning first to FIG. 1, there is shown a schematic perspectiveview of the osmometer 10 of the present invention. The osmometer 10includes a sample cell 12. Sample cell 12 includes a chamber body 14which is preferably made of polysulfone and is preferably transparent.An Amicon 8400 stirred cell dialysis chamber serves well as sample cell12. Residing in chamber body 14 is membrane 16. Many semi-permeablemembranes are available commercially which work well. For aqueoussolutions a Diaflo ultrafiltration membrane YM10 (10K Mw cut-off) waseffectively used for membrane 16. Other membranes could be chosen withtighter (or looser) pores to selectively measure the osmoticcontribution of lower (or higher) molecular weight fractions of thesample solution. Many of the commercially available membranes are suitable for organic solvents. The Diaflo YM10 membrane is suitable for mostorganic solvents as well, excluding Amines, phenols and solutions withpH less than 3 or greater than 13. This is not a limitation of thedevice; other membranes are commercially available which circumvent allof these limitations. Membrane 16 is supported on meandering dialyzatecell 18. Meandering dialyzate cell 18 is retained in chamber body 14 bymeans of base plate 20 which threadably engages chamber body 14. AnO-ring 22 provides a seal between chamber body 14 and meanderingdialyzate cell 18. There is a circumferential lip 24 in the interiorsurface of chamber body 14. Circumferential lip 24 provides residencefor support bracket 26 which preferably includes three radial spokes 28.Extending down from support bracket 26 is stir rod axle 29. Rotatablymounted on stir rod axle 29 is stir rod blade 30.

[0023] Press fit onto the top of chamber body 14 is lid 32. A sealbetween lid 32 and chamber body 14 is provided by means of O-ring 35.Attached to lid 32 is bushing 34 which aligns with bore 36 in lid 32.Extending from bushing 34 is pressurized gas conduit 38 for whichpressurized gas is supplied from a pressurized gas source 40. Mounted inpressurized gas conduit 38 is a pressure regulator 42 and a pressuregauge 44. Lid 32 is also provided with an L-shaped bore 46 in which apressure relief valve 48 is mounted. Pressure relief valve 48 ismanually operated by means of handle 50.

[0024] There is a bore 52 into meandering dialyzate cell 18 whichcommunicates with one of radial channels 55 (see FIG. 2) in the topsurface of meandering dialyzate cell 18. The top surface of meanderingdialyzate cell 18 also includes a series of concentric channels 57therein. Bore 52 aligns with bore 54 through chamber body 14. Coupling56 mounts to chamber body 14 at bore 54 and transparent dialyzate exittube 58 extends therefrom.

[0025] When in operation, sample cell 12 resides in frame 60 (shown in aperspective view in FIG. 3) which is preferably open on at least twosides thereof to permit observation of sample cell 12. Frame 60 is madeof metal (preferably steel) and insures that lid 32 is retained onchamber body 14 when sample cell 12 is pressurized via pressurized gasconduit 38. Frame 60 is supported on magnetic stirring/hot plate 62. Anexemplary magnetic stirring/hot plate 62 that may be used in thepractice of the present invention is a Coming Hot Plat/Stirrer modelPC-351. Because frame 60 is metallic, it acts as a good heat conductorfor heating sample cell 12.

[0026] Initially the pressurizing lid 32 is removed and the samplesolution 64 is introduced into the chamber 14 above the membrane 16. Thegas delivered via pressurized gas conduit 38 can be air, nitrogen or anon-interacting (inert) gas. Preferably, pressurized gas source 40 candeliver gas at a relatively high pressure (80 psi). The air pressureapplied to the sample solution 64 is controlled by the pressureregulator 42 which has the capability of smoothly varying the pressureover the entire desired range of measurement (0-10 psi for low pressureapplications, 0.2-75 psi for high pressure applications). Pressureregulators with more or less sensitivity can be chosen based upon theosmotic pressure of the sample solution being measured and the desiredaccuracy. Two examples of pressure gauges can be used in the operationof the present invention are the Nullmatic 40-30 pressure regulator andthe Ashcroft 40 psi pressure regulator. The applied pressure is measuredon pressure gauge 44. The accuracy and range of the osmometer 10 dependson the accuracy and pressure range of the pressure gauge 44 selected. ACapsuhelic 0-300 inch of water gauge has been used successfully forlower pressure solutions. Additional accuracy can easily be obtained byusing any of a number of more expensive and accurate commerciallyavailable pressure gauges.

[0027] The sample cell 12 plus solution 64 is weighed and then the lid32 is sealed with the pressure release valve 48 open. The sealed samplecell 12 is then placed inside the metal pressure frame 60 and thepressure release valve 48 is closed. This frame 60 holds the lid 32firmly in place under pressurization. The base of the frame 60establishes good thermal contact between the magnetic stirring/hot plate62 and the sample cell 12. The heating element of the magneticstirring/hot plate 62 is turned on and set to the chosen temperaturesetting. The entire apparatus can be submerged into a water bath (notshown) with a temperature sensor (not shown) for precise temperaturecontrol. Because of the large volume of sample solution 64 and the fastmeasurement time this step is often not needed in the practice of themethod of present invention. The magnetic stir portion of the magneticstirring/hot plate 62 is turned on thereby magnetically driving theangled magnetic stirring rod blade 30 which is preferably set to amoderate rate of stirring of about 1-2 revolutions per second. For clearsolutions the surface of the membrane 16 is then visually inspectedthrough the transparent sides of the chamber 14 for bubbles or debris.

[0028] The pressure is raised initially to between 5 and 15 psi to wetthe membrane 16 with the sample solution 64. Once the sample solution 64is forced through the membrane 16 and the dialyzate begins to emergethrough the transparent dialyzate exit tube 58, such that there is avisible meniscus 66 therein, the pressure is reduced using the pressureregulator 42 until flow ceases. Pressure is reduced further until flowreverses direction and the dialyzate is drawn back into the meanderingdialyzate cell 18 and ultimately back through the membrane 16 into thetransparent sample chamber 14. The meandering channel dialyzate cell 18maximizes the surface area for equilibration while minimizing the volumeof dialyzate and thus the amount of dilution of the sample 64. This is adramatic improvement over the commercial membrane osmometers that havemeandering channel sample chambers which minimize the sample volumerather than the dialyzate volume. For fast and easy use a minimumdialyzate volume to sample volume ratio is preferable. For expensivesamples a smaller cell can be selected which maintains the large samplevolume to dialyzate volume ratio but reduces the dimensions of theentire cell. Finally, the pressure is varied carefully until themeniscus 66 in the dialyzate exit tube 58 holds substantiallystationary, that is, stationary over a few minute time period. Theosmotic pressure of the sample solution 64 is equal to the applied gaspressure read upon the pressure gauge 44 when the flow is substantiallystationary, that is when equilibrium across the membrane is reached.Preferably, the osmotic pressure measured is then corrected for theslight hydrostatic pressure difference calculated from the difference inheight of the liquid column in the dialyzate exit tube 58, and theheight of the sample surface 64 in the sample cell 12 (typically thiscorrection is between 1 and 10 centimeters of water). This technique ofmeasuring the hydrostatic pressure differential can be adapted into amethod for achieving increased accuracy, particularly in low pressureapplications (0-1 psi). Increased accuracy in low pressure applicationscan be accomplished by suspending the dialyzate tube vertically andmeasuring the difference in heights of the stationary meniscus of thedialyzate tube 66, and the height of the sample surface 64 in the samplecell 12. The osmotic pressure is then calculated by correcting the gaugepressure for the hydrostatic pressure difference. Preferably, the stepof measuring the difference in heights of the stationary meniscus 66 inthe dialyzate tube 58, and the height of the sample surface 64 in thesample cell 12 is performed at two or three applied pressures typicallydiffering by a few centimeters of water (1-5cm). The osmotic pressure isthen calculated by correcting the gauge pressure for the hydrostaticpressure difference for each chosen pressure. The two or three osmoticpressure values obtained are then averaged. Typically the pressureschosen are only different by a few centimeters of water to avoid largecolumns of dialyzate in the vertical dialyzate exit tube which wouldrequire large dialyzate volumes.

[0029] For measuring a dilution series the pressure relief valve 48 isopen, the lid 32 is removed, the diluent is added and the sample isreweighed so that the new concentration is known. The lid 32 isreaffixed and put in the metallic frame 60. Finally the pressure reliefvalve 48 is closed and the measurement method is repeated as describedabove. This entire process can be performed in just a few minutes.

[0030] An alternate method of measuring the osmotic pressure of a samplesolution is to set the pressurized gas conduit 38 to a fixed pressureusing the pressure regulator 42 and pressure gauge 44 and then wait forthe solution to dialyze down to the concentration corresponding to thispressure. The solution can then be removed and its concentration can bedetermined using other techniques (spectrophotometrically orgravimetrically for instance). This mode of operation allows for thepreparation of solutions of known osmotic pressure. This is not thepreferred mode of use because the dialysis process is quite timeconsuming, sometimes taking several hours to reach the chosen osmoticpressure.

[0031] Looking next at FIG. 4, the accuracy of the present invention isdemonstrated using some standard solutions from the prior artliterature. Clearly the data generated using the method and apparatus ofthe present invention is in good agreement with the polystyrenesulfonate data measured using conventional membrane osmometry. Further,the highest concentration point obtained using the invention was beyondthe viscosity and pressure range accessible using conventional membraneosmometry methods.

[0032] Turning to FIG. 5, it can be seen that the osmotic pressure datawas obtained for three complex solutions using the present invention.The osmotic pressure of these three complex solutions cannot be measuredusing prior art membrane osmometry techniques. The first solution is a143 centipoise charged polymer solution made from mixing gelatin and asynthetic polyelectrolyte. This high viscosity solution cannot bemeasured using the meandering sample channel arrangement of commercialmembrane osmometers because of the high backpressure needed to introducethe solution. In addition, this high osmotic pressure is slightly beyondthe nominal operating limits of commercial membrane osmometers (althoughmost transducers are still fairly linear in this range). The secondsolution is a carbon black slurry dispersed in gelatin. While the highmeasured osmotic pressure of this solution is slightly beyond theoperating limits of conventional membrane osmometers, the moresignificant problem with this solution is that the introduction of sucha carbon black slurry into commercial osmometers will foul the osmometerseverely. The procedure for cleaning and recalibration of the osmometerafter this solution requires several hours and makes these sorts ofmeasurements time prohibitive as a practical matter. In contrast, in theoperation of the present invention another solution can be measuredwithin minutes. The third solution measured is a yellow photographic(silver halide) dispersion. This solution fouls conventional membraneosmometers and gives data which is not reproducible because of settlingand surface plugging. These problems are avoided using the presentinvention because of the constant stirring of the solution and sweepingof the surface.

[0033] As discussed above, those skilled in the art will recognize thatthe present invention has many advantages over prior art osmometers. Forexample, it should be appreciated that the present invention is easilydismantled and cleaned so that complex fluids (emulsions anddispersions) can be measured which would foul traditional meanderingchannel membrane osmometers. In addition, the magnetic stirring rodblade 30 that continually sweeps the surface of the membrane 16, reducessurface fouling and removes bubbles that hamper the accuracy andreliability of commercial prior art membrane osmometers when used tomeasure complex fluids. Further, because chamber body 14 of the deviceis transparent, the upper surface of the membrane 16 may be observed tocheck for detrimental bubbles, deposits, etc. In addition, the magneticstirrer keeps the sample well mixed, avoiding particle settling and thedevelopment of surface concentration gradients which can lead toinaccurate measurements in commercial membrane osmometers of the priorart when used with complex fluids (particularly dispersions).

[0034] Sample measurements in the present invention are very fast,approximately 4-50 times faster than other prior art methods. Because ofthe speed of use of the present invention it can be used to measure theosmotic pressure for solutions that change as a function of time ortemperature. In fact, chemical reactions can be initiated in theosmometer sample cell and then monitored as a function of time.Reactants can be added to the sample cell at the desired temperature,and a timer can be started. The sample cell 12 is sealed and placed onthe magnetic stirring/hot plate 62 where the temperature is maintainedand mixing is initiated. The sample cell 12 is pressurized and thepressure is varied until the dialyzate meniscus 66 in the exit tube 58is substantially stationary, that is, there is substantial equilibriumacross the membrane. The meniscus 66 may not become entirely stationaryif the osmotic pressure of the reacting sample is changing too rapidly.The pressure, as read from pressure gauge 44, required to keep themeniscus 66 stationary, is then recorded as a function of time. Thesemeasurements can be recorded repeatedly at a time step as small as 1-2minutes to monitor the evolution of a chemical (or biological) reactionthat changes osmotic pressure with time. Chemical and biologicalreactions that change osmotic pressure with time include an enormousclass of reactions that use up or produce reactants.

[0035] An alternative embodiment of the method of the present inventionis to monitor the changes in osmotic pressure of a sample or ongoingchemical reaction as a function of time as the temperature is varied.This involves using a hot plate or water bath capable of varying thetemperature of the sample. To measure the temperature one can affix athermometer 70 or a thermocouple probe or other temperature sensingdevices to the cell. In this case the sample (or reactants) are added atsome prescribed temperature to the sample cell 12, and a timer isstarted. The sample cell 12 is sealed and placed on the magneticstirring/hot plate 62 (or in the water bath) where mixing is initiatedin the sample cell 12 and the temperature is varied in somepredetermined way as a function of time. Examples include ramping thetemperature either upward or downward, or causing step changes in thetemperature of the sample 64. The sample cell 12 is pressurized and thepressure is varied until the dialyzate meniscus 66 in the exit tube 58is nearly stationary. The meniscus 66 may not become entirely stationaryif the osmotic pressure of the sample 64 is changing too rapidly. Thepressure on the gauge 44 needed to keep the meniscus 66 stationary isthen recorded as a function of time and temperature. This is the osmoticpressure. These measurements can be recorded repeatedly at a time stepas small as 1-2 minutes to monitor the evolution of a chemical (orbiological) reaction that changes osmotic pressure with temperature andtime. These techniques would be particularly suitable for monitoring theprogress of chemical or physical gelation reactions for example. Theosmometer 10 of the present invention can clearly be used to measure theosmotic pressure of samples 64 which are unstable if held more than 5-10minutes (for example due to chemical or colloidal instabilities) orchange substantially within 5-10 minutes. Previously these types ofmeasurements simply were not possible with commercial prior art membraneosmometers. This alternative embodiment can be used to monitor thechanges in osmotic pressure of a sample as a function of time as thetemperature is varied, not only for chemical reactions, but also forthose changes in osmotic pressure resulting From, for example, phasechanges, morphological changes, colloidal instabilities including Oswaldripening, aggregation, counter ion condensation, and micelle formation.

[0036] Sample measurements in the device of the present invention arevery fast, approximately 4-50 times faster than other prior art methods.Because of the speed of use of the present invention it can be used tomeasure the osmotic pressure for solutions that are unstable if heldmore than 5-10 minutes (for example due to chemical or colloidalinstabilities). Previously these measurements simply were not possiblewith commercial prior art membrane osmometers. The present invention isfaster because the sample volume is typically several hundred timeslarger than the dialyzate volume so that the change in concentrationupon equilibration can be safely ignored allowing accurate measurementsto be made immediately (1-5 minutes per sample). In contrast, typicalprior art membrane osmometers require multiple flushes of each samplesolution before concentrations stop changing and reproducibility isestablished (˜20-50 minutes per sample). A second reason that the deviceof the present invention allows for faster measurements is that newsolutions can be mixed directly in the sample cell 12. Accurate dilutionseries are made by starting from a sample solution 64 and progressivelyadding known amounts of diluent. This is achieved practically by simplyplacing the entire cell on an accurate balance and weighing the amountof added solution before each measurement. This sort of mixing andweighing is not possible in conventional art osmometers and thusrequires the individual preparation of separate samples for dilutionseries. In addition, because the sample cell 12 is not mixed inconventional prior art osmometers, between each measured dilutionseveral extra flushes are typically required to achieve stablemeasurements, further slowing down the measurement process. In theoperation of the present invention, calibration does not requireequilibration of a sample. It is simple and fast based on accuratelymeasuring the air-line pressure. This is in marked contrast to thedifficulty of repeatedly calibrating transducers and properly seatingmembranes in commercial prior art membrane osmometers. This leads tosignificant time savings in operation.

[0037] Further, the accuracy of the device of the present invention issignificantly greater than with prior art osmometers. The accuracy ofthe device of the present invention is determined by the air pressuregauge which can be controlled very precisely (to within millimeters ofwater if desired). Commercial prior art osmometers can only achieve 1-2cm of water accuracy. The applicable pressure range which can be used inthe operation of the present invention is determined by the sample cellmaximum (which can withstand up to 75 psi), far higher than theconventional transducer technology used in prior art membrane osmometerswhich typically measure in a range of 1.5-3 psi).

[0038] Finally, the present invention enables the determination of theosmotic pressure of high viscosity solutions. Meandering channelosmometers of the prior art have small sample volumes and small samplechannels and due to high backpressure are unusable with high viscositysolutions (above about 100 cp).

[0039] In the preferred embodiment, transparent dialyzate exit tube 58has been described herein as extending from a coupling 56 attached tothe chamber body 14 at bore 54. Those skilled in the art will recognizethat a variety of different structural arrangements are available forproviding and attaching the transparent dialyzate exit tube 58.Transparent dialyzate exit tube 58 can even originate inside chamberbody 14 so long as the meniscus can still be viewed. As such, dialyzateexit tube 58 will be attached, either directly or indirectly, to someportion of sample cell 12. The dialyzate exit tube 58 must, however, betransparent and should be oriented at any angle from horizontal to aconfiguration where a large portion thereof is oriented verticallyupward. By vertically upward, it is meant that the dialyzate exit tube58 can be oriented vertically with the end thereof which is open toatmosphere residing at a higher elevation than the remainder of exittube 58.

[0040] In the description of the method and alternative methods of thepresent invention, the sample or solution is described as beingintroduced to the sample cell 12 above the membrane 16. Those skilled inthe art will recognize that a variety of configurations are availablefor sample cell 12 which may not be in agreement with the traditionalmeaning of the terms “above” and “below”. For example, the sample cellmay take the shape of a U-shaped cylinder with the membrane positionedperpendicular to the cylindrical axis of the U-shaped cylinder almostanywhere along the length thereof. With that in mind, it is appropriateto think of the sample cell 12 being divided by the membrane 16 suchthat there is a sample side of the membrane 16, (that side of themembrane 16 where the sample or solution is added to the sample cell12), and a dialyzate side of the membrane16. For the purposes of thisapplication “above” is intended to mean on the sample side of themembrane 16.

[0041] The present invention has far reaching potential uses. It allowsfor easy measurement of osmotic pressure of complex fluids and could beused as a method for screening drugs for activity in osmoticallyimplicated biological phenomena such as blood pressure changes,arthritic swelling of joints and excessive ocular pressure. It makespossible the study of the evolution polymerization reactions, andcolloidal instability phenomenon. The osmotic pressure gives a directmeasure of the appearance or disappearance of macromolecular chargedspecies. This sort of research has been hampered by the problems ofcommercial prior art osmometers overcome by this invention.

[0042] From the foregoing, it will be seen that this invention is onewell adapted to obtain all of the ends and objects hereinabove set forthtogether with other advantages which are apparent and which are inherentto the invention.

[0043] It will be understood that certain features and subcombinationsare of utility and may be employed with reference to other features andsubcombinations. This is contemplated by and is within the scope of theclaims.

[0044] As many possible embodiments may be made of the invention withoutdeparting from the scope thereof, it is to be understood that all matterherein set forth and shown in the accompanying drawings is to beinterpreted as illustrative and not in a limiting sense. PARTS LIST: 10osmometer 12 sample cell 14 chamber body 16 membrane 18 meanderingdialyzate 20 base plate 22 o-ring 24 circumferential lip 26 supportbracket 28 radial spokes 29 stirrod axle 30 stir rod blade 32 lid 34bushing 35 o-ring 36 bore 38 pressurized gas conduit 40 pressurized gassource 42 pressure regulator 44 pressure gauge 46 L-shaped bore 48pressure relief valve 50 handle 52 bore 54 bore 55 radial channels 56coupling 57 concentric channels 58 transparent dialyzate exit tube 60frame 62 magnetic stirring/hot plate 64 sample solution 66 meniscus 70thermometer

What is claimed is:
 1. A method for measuring osmotic pressure of asolution comprising the steps of: (a) placing a sample of a solutioninto a sample cell above a membrane supported in the sample cell; (b)sealing the sample in the sample cell; (c) introducing gas at anelevated pressure to the sample cell to drive a quantity of dialyzatethrough the membrane and into a transparent dialyzate exit tube adaptedto receive the dialyzate which passes through the membrane; and (d)varying the elevated pressure of the gas to yield a substantiallystationary dialyzate meniscus in the transparent dialyzate exit tube. 2.A method as recited in claim 1 further comprising the step of: readingthe pressure gauge showing the elevated pressure of the gas that yieldedthe substantially stationary dialyzate meniscus, the elevated pressureof the gas being substantially equivalent to the osmotic pressure of thesample.
 3. A method as recited in claim 1 further comprising the stepof: reducing the elevated pressure of the gas to minimize the volume ofdialyzate in the dialyzate exit tube prior to said varying step.
 4. Amethod as recited in claim 1 further comprising the step of: maintainingthe sample in the sample cell at a predetermined temperature.
 5. Amethod as recited in claim 1 further comprising the step of: continuallystirring the sample in the sample cell.
 6. A method as recited in claim2 further comprising the step of: performing a hydrostatic pressurecorrection based on the height of the solution in the sample cell in theheight of the dialyzate in the dialyzate exit tube.
 7. A method formeasuring osmotic pressure of a solution comprising the steps of: (a)placing a sample of a solution into a sample cell, the sample cellincluding therein a membrane supported on a meandering dialyzate cell;(b) sealing the sample in the sample cell; (c) introducing gas at anelevated pressure to the sample cell to drive a quantity of dialyzatethrough the membrane and the meandering dialyzate cell to enter adialyzate exit tube extending from the sample cell and obtaining astationary dialyzate meniscus in the dialyzate exit tube; (d) measuringthe difference in elevation of the sample in the sample cell and themeniscus in point to the dialyzate exit tube; (e) reading a finalpressure on the pressure gauge showing the elevated pressure of the gasthat yielded the stationary dialyzate meniscus; and (f) calculating theosmotic pressure from the final pressure and the difference inelevation.
 8. A method as recited in claim 2 further comprising the stepof: (a) opening the sample cell and adding additional sample solutionthereto; (b) weighing the sample cell with the additional samplesolution therein; and (c) repeating steps (b) through (d) of claim
 1. 9.A method as recited in claim 7 further comprising the step of: (a)opening the sample cell and adding additional sample solution thereto;(b) weighing the sample cell with the additional sample solutiontherein; and (c) repeating steps (b) through (f) of claim
 7. 10. Amethod as recited in claim 1 further comprising the step of: collectingthe dialyzate passing through the membrane in a dialyzate collector andflowing the dialyzate therefrom to the transparent dialyzate exit tube.11. A method as recited in claim 1 further comprising the step of:collecting the dialyzate passing through the membrane in a meanderingdialyzate cell and flowing the dialyzate therefrom to the transparentdialyzate exit tube.
 12. A method as recited in claim 2 furthercomprising the step of: reading the pressure gauge at predeterminedintervals after repeating said varying step to determine the change inosmotic pressure of the solution as a function of time.
 13. A method asrecited in claim 11 further comprising the step of: changing thetemperature as a function of time during said reading step.
 14. A methodfor measuring osmotic pressure of a solution comprising the steps of:(a) placing a sample of a solution into a sample cell above a membranesupported in the sample cell; (b) sealing the sample in the sample cell;(c) introducing gas at an elevated pressure to the sample cell togenerate a dialyzate by allowing a portion of the solution to dialyzethrough the membrane; and (d) varying the elevated pressure of the gasuntil equilibrium is achieved and no more dialyzate passes the membrane.15. An apparatus for measuring osmotic pressure of a solutioncomprising: (a) a sample cell for receiving a sample of a solution, saidsample cell including a chamber with a meandering dialyzate cell thereinand a membrane supported in said sample cell; (b) a removable lidsealingly engagable with said sample cell; (c) a gas inlet port throughwhich gas at an elevated pressure is delivered to said sample cell tocause a portion of a sample solution in said sample cell to dialyzethrough said membrane and enter a transparent dialyzate exit tube; (d)an inlet conduit connected to said gas inlet port, said inlet line alsobeing connected to a source of pressurized gas; (e) a pressure regulatorconnected to said inlet conduit between said source of pressurized gasand said gas inlet port; and (f) a pressure gauge connected to saidinlet conduit between said pressure regulator and said gas inlet port.16. An apparatus as recited in claim 15 further comprising: means formaintaining the sample in the sample cell at a predeterminedtemperature.
 17. An apparatus as recited in claim 15 further comprising:a magnetically driven stirring blade in said sample cell above saidmembrane.
 18. An apparatus as recited in claim 15 further comprising: arotating stirring blade in said sample cell above said membrane.
 19. Anapparatus as recited in claim 15 further comprising: a heater formaintaining the sample in the sample cell at a predeterminedtemperature.
 20. An apparatus as recited in claim 15 further comprising:a dialyzate collector positioned beneath said membrane.
 21. An apparatusas recited in claim 15 wherein: said dialyzate collector is a meanderingdialyzate cell.
 22. An apparatus as recited in claim 20 wherein: saidtransparent dialyzate exit tube receives dialyzate from said dialyzatecollector.
 23. An apparatus as recited in claim 15 wherein: said samplecell includes a transparent chamber body and a base plate which isdetachable from and re-engagable to said chamber body.